Leica BIOSYSTEMS DS9477 Mode D'emploi page 2

ChromoPlex 1 Dual Detection for BOND
Catalog No: DS9477
Intended Use
This detection system is for in vitro diagnostic use.
ChromoPlex 1 Dual Detection is a biotin-free, polymeric horseradish peroxidase (HRP)-linker and polymeric alkaline phosphatase
(AP)-linker antibody conjugate system for the detection of tissue-bound mouse and rabbit IgG primary antibodies. It is intended for
staining sections of formalin-fixed, paraffin-embedded tissue on the BOND automated system (includes Leica BOND-MAX system and
Leica BOND-III system).
The clinical interpretation of any staining or its absence should be complemented by morphological studies. Proper controls should be
evaluated within the context of the patient's clinical history and other diagnostic tests by a qualified pathologist.
The ChromoPlex 1 Dual Detection must be used with laboratory best practice in the use of tissue controls. For assurance, laboratories
should stain each patient sample in conjunction with positive, negative and other tissue specific controls as needed.
Summary and Explanation
Immunohistochemical techniques can be used to demonstrate the presence of antigens in tissue and cells (see "Using BOND Reagents"
in your BOND user documentation).
ChromoPlex 1 Dual Detection utilizes a novel controlled polymerization technology to prepare polymeric HRP-linker and AP-linker
antibody conjugates. The detection system avoids the use of streptavidin and biotin, and therefore eliminates non-specific staining as a
result of endogenous biotin.
ChromoPlex 1 Dual Detection works as follows:
• The specimen is incubated with hydrogen peroxide to quench endogenous peroxidase activity.
• A user-supplied specific primary mouse and rabbit antibody cocktail is applied.
• Poly-HRP IgG reagent localizes mouse antibodies.
• Poly-AP IgG reagent localizes rabbit antibodies.
• The first substrate chromogen, 3,3'-Diaminobenizidine tetrahydrochloride hydrate (DAB), visualizes mouse antibodies via a brown
precipitate.
• The second substrate chromogen (Fast Red) visualizes rabbit antibodies via a red precipitate.
• Hematoxylin (blue) counterstaining allows the visualization of cell nuclei.
Using ChromoPlex 1 Dual Detection in combination with the BOND automated system reduces the possibility of human error and
inherent variability resulting from individual reagent dilution, manual pipetting and reagent application.
Reagents Provided
The reagents provided are sufficient for 25 individual BOND staining runs, or a maximum of 100 slides.
To achieve a maximum of 100 slides from this detection system, slides must be batched in quantities of 4 or greater, per Slide Staining
Assembly. Batching in quantities of less than 4 will result in fewer stained slides.
1. Peroxide Block (15 mL) 3-4% Hydrogen peroxide.
2. Polymer mHRP (15 mL) Poly-HRP anti-mouse containing 10% (v/v) animal serum in Tris-buffered saline and 0.1%
3. Polymer rAP (15 mL) Poly-AP anti-rabbit containing 10% (v/v) animal serum in Tris-buffered saline and 0.09%
4. DAB Part 1 (2 mL) 66 mM 3,3'-Diaminobenzidine tetrahydrochloride hydrate in a stabilizer solution.
5. DAB Part B (35 mL) ≤0.1% (v/v) Hydrogen peroxide in a stabilizer solution.
6. Red Part A (5.5 mL) Activator containing 0.5% ProClin™ 950.
7. Red Part B (1.0 mL) Substrate.
8. Red Part C (1.0 mL) Substrate.
9. Red Part D (35 mL) Buffer solution containing 0.5% ProClin™ 950.
10. Hematoxylin DS9477 (15 mL) <0.1% Hematoxylin.
Dilution and Mixing
ChromoPlex 1 Dual Detection is optimized for use on the BOND system.
Reconstitution, mixing, dilution, or titration of these reagents is not required.
Materials Required But Not Provided
Mounting media compatible with ChromoPlex 1 Dual Detection (see Precautions).
Refer to "Using BOND Reagents" in your BOND user documentation for a complete list of materials required for specimen treatment and
immunohistochemical staining using the BOND system.
Storage and Stability
Store at 2–8 °C. Do not freeze. Do not use after the expiration date indicated on the tray handle label. Return to 2–8 °C immediately after
use.
There are no obvious signs to indicate instability of this product; therefore positive and negative controls should be run simultaneously
with unknown specimens (refer to "Quality Control" in the "Using BOND Reagents" section of your BOND user documentation).
If unexpected staining is observed that cannot be explained by variations in laboratory procedures, and a problem with the detection
system is suspected, contact your local distributor or the regional office of Leica Biosystems immediately.
Storage conditions other than those specified above must be verified by the user
DS9477
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ProClin™ 950.
ProClin™ 950.